三叶青提取物诱导肺癌A549株细胞凋亡的研究

目的探讨三叶青提取物对肺癌细胞A549的作用,初步探讨其机制。方法采用体外细胞培养技术,采用流式细胞仪检测A549细胞株的细胞周期分布相和细胞凋亡率。结果流式细胞仪检测结果显示:加三叶青提取物10-1g/L、10-2g/L处理后的细胞凋亡比例明显高于对照组。结论三叶青提取物对A549细胞具有诱导凋亡的作用。     

虎杖粗提物对人肺癌A549细胞株诱导凋亡作用的形态学观察

应用MTT法检测虎杖提取物对体外培养的人肺癌A549细胞株的抑制增殖作用的影响,通过倒置显微镜、HE染色、AO/EB荧光显微镜的方法观察肿瘤细胞的形态改变.结果 显示,虎杖提取物在体外对A549人肺癌细胞株有明显的抑制增殖作用,而且这种抑制作用呈现浓度和时间的依赖性.细胞的形态学观察发现,虎杖提取物作用后出现凋亡细胞.认为虎杖提取物在体外对人肺癌A549细胞株有显著的抑制增殖和诱导凋亡作用.     

山茱萸水提取物对人肺癌A549细胞凋亡的作用

目的探讨山茱萸水提取物对人肺癌A549细胞的体外增殖的抑制及凋亡作用。方法采用MTT法及DNA琼脂糖凝胶电泳法检测山茱萸水提取物对A549细胞增殖的抑制及凋亡作用,并观察细胞形态改变。结果山茱萸水提取物对A549细胞抑制作用较明显,在一定范围内呈现较好的量效、时效关系;经不同剂量的山茱萸水提取物处理的细胞电泳均出现相差约200 bp的DNA梯状电泳图谱,形态学观察到细胞凋亡。结论山茱萸水提取物对人肺癌A549的细胞增殖有较强的抑制作用,能诱导其细胞凋亡。     

蛴螬提取物对人肺癌A549细胞Bax和p21蛋白表达的影响

目的观察蛴螬提取物对人肺癌A549细胞增殖的影响及诱导凋亡的机制。方法采用MTT法检测蛴螬提取物对人肺癌A549细胞的增殖抑制率;运用免疫细胞化学SP法检测用药前后Bax和p21蛋白表达的变化;运用流式细胞仪检测细胞周期及细胞凋亡率。结果蛴螬提取物对人肺癌A549细胞具有明显的增殖抑制作用,并呈时间依赖性;蛴螬提取物组细胞Bax和p21表达均增强,细胞凋亡率与对照组相比有显著性差异,并被阻滞在细胞周期的S期。结论蛴螬提取物可通过上调Bax和p21使细胞阻滞于S期,从而抑制A549细胞的生长。     

藤梨根乙酸乙酯提取物对肺癌A549细胞凋亡的诱导作用

目的观察藤梨根乙酸乙酯提取物对肺癌A549细胞凋亡的诱导作用。方法体外培养肺癌A549细胞,分别给予40、80、160μg/ml的藤梨根乙酸乙酯提取物（实验组）,同时设对照组（0.04%DMSO＋肺癌A549细胞）。采用DNA琼脂糖凝胶电泳检测用药后DNA梯状条带、TUNEL检测用药后肺癌A549细胞凋亡率的变化、免疫组化SP法检测用药后肺癌A549细胞Survivin蛋白表达变化、RT-PCR法检测用药后A549细胞Survivin mRNA的表达。结果实验组细胞DNA凝胶电泳均出现凋亡细胞所特有的DNA梯状条带图谱,而对照组细胞未出现。各实验组细胞随着藤梨根乙酸乙酯提取物浓度的增加,细胞凋亡率明显增加,并存在时相性和量—效依赖性;其Survivin蛋白和mRNA表达均低于对照组（P均〈0.05）,亦存在时相性和量—效依赖性。结论藤梨根乙酸乙酯提取物可明显诱导肺癌A549细胞凋亡,其机制可能与降低Survivin蛋白和mRNA表达有关。     

微波对人肺癌A549细胞凋亡的非热效应

目的了解微波对人肺癌A549细胞的非热作用。方法在冰浴排除热效应条件下用不同剂量微波(12 mV/cm2、18 mV/cm2和21mV/cm2)照射A549细胞10 min,24 h光镜观察照相后收集细胞,用超声裂解法提取蛋白,Western印迹检测凋亡相关蛋白表达变化;Rh123染色行流式分析细胞线粒体膜电势;间接免疫荧光法检测凋亡蛋白活化的caspase3变化;Hoechst荧光染色检测细胞凋亡。结果①光镜下所见:与对照组相比,照射强度越大细胞数减少越明显,细胞质越致密;②Rh123染色流式分析:随着微波照射强度的增加,线粒体膜电势降低,高荧光密度群百分率减少,荧光密度平均群百分率减少下降、低荧光密度群百分率增加(2.9%vs 5.4%、7.6%、9.9%;各照射组均高于对照组,P<0.01);③Hoechst荧光染色检测到微波辐射导致A549细胞凋亡,照射强度越大,细胞凋亡越多;④间接免疫荧光法检测到微波辐射导致A549细胞凋亡蛋白Caspase3活化增多,随照射强度升高表达明显增加;⑤Western印迹:各照射组凋亡相关蛋白Bax/Bcl-2、Caspase3均高于对照组(P<0.01),照射强度越大此值越高。结论微波对人肺癌A549细胞存在明显的促凋亡非热效应,且与照射强度呈正相关。     

白藜芦醇诱导肺腺癌A549细胞凋亡的机制

目的探讨白藜芦醇(Res)诱导肺腺癌A549细胞凋亡的机制。方法用不同浓度Res处理A549细胞24 h后,倒置显微镜观察细胞形态学变化,4,6-二氨基-2-苯基吲哚(DAPI)细胞核染色后荧光显微镜观察细胞凋亡特征,噻唑蓝(MTT)检测对于A549细胞的抑制生长作用,Western印迹法检测Res作用A549细胞后P53、Bax、Bcl-2、Cleaved-Caspase3蛋白的表达变化。结果 Res处理A549细胞后,细胞间隙变大,细胞核分裂,随着药物浓度的增加上述形态变化明显。25²00μmol/L浓度的Res可明显抑制A549细胞的增殖,具有剂量依赖性,24 h的IC50为100μmol/L,而且细胞内的P53、Bax、Cleaved-Caspase3蛋白表达量升高,Bcl-2蛋白被抑制,Bcl-2/Bax比值下降。结论 Res抑制A549细胞增殖,并通过P53途径诱导A549细胞凋亡,Bax、Bcl-2和Cleaved-Caspase3参与了凋亡过程。     

微波辐射诱导肺癌A549细胞凋亡的研究

微波能诱发肿瘤细胞凋亡，故能在微波凝固去除原发灶的同时抑制肿瘤邻近扩散，这对沿气道管壁或管外浸润生长的肿瘤、纤维支气管镜清除术后的进一步局部干预很有意义。本研究旨在探讨不同功率和作用时间的微波辐照后对肺癌A549细胞凋亡的影响及机制。[第一段]     

山茱萸多糖对肺癌A549细胞凋亡的影响及机制

目的探讨山茱萸多糖对人肺癌A549细胞凋亡作用的影响及机制。方法收集对数生长期A549细胞,加入终质量浓度分别为25、50、100 mg/mL的山茱萸多糖20μL,分别培养24、48、72 h,MTT法测定细胞增殖抑制率和山茱萸多糖的半抑制浓度(IC50)。倒置显微镜下观察50 mg/mL山茱萸多糖作用后细胞形态变化。免疫组化SP法检测Survivin蛋白表达,用免疫组化评分(IHS)表示。结果山茱萸多糖作用后A549细胞生长受到明显抑制,细胞增殖抑制率呈明显的量效、时效关系;倒置显微镜下观察到细胞凋亡的典型改变,免疫组化SP法染色显示Survivin表达降低,Survivin蛋白的IHS随浓度增加呈明显下降趋势。结论山茱萸多糖能诱导A549细胞凋亡,其机制可能与下调Survivin表达有关。     

槐耳清膏诱导人肺腺癌细胞A549凋亡的实验研究

槐耳清膏是药用真菌槐耳菌质的热水提取物 ,临床上证实有一定的抗癌效果。为进一步探讨其抗癌机制 ,我们研究了槐耳清膏在体外诱导人肺腺癌细胞系Ａ5 49凋亡的作用。材料与方法　引进并培养人肺腺癌细胞系Ａ5 49细胞株。制备不同浓度的槐耳清膏ＰＲＭＩ 16 40含药培养液与接种成功、呈对数生长的Ａ5 49细胞共同培养 2 4ｈ后 ,每 6ｈ进行以下观察及测定 (设阳性对照组羟基喜树碱 10 0 μｇ/ｍｌ,共同孵育 10～ 12ｈ后进行测定 ) [1] 。 (1)荧光染色 :收集培养细胞 ,清洗并均匀涂片自然风干 2 4ｈ ,置于 0 0 5ｍｇ/Ｌ的Ｈｏｅｃｈｓｔ332 5 8…     

Safrole oxide induces apoptosis in A549 human lung cancer cells

3,4-(Methylenedioxy)-1-(2',3'-epoxypropyl)-benzene (safrole oxide) was synthesized in the authors' laboratory. To investigate the effects of safrole oxide on the growth and apoptosis of A549 human lung cancer cells, the authors treated the cells with safrole oxide, 112.36 to 449.44 micromol/L, for 24 to 48 hours. The results showed that the drug led A549 cells to apoptosis and blocked cell cycle completely at G1 phase and partly at G(2)-M phase. To further study the correlated mechanism, the authors examined P53 and H-Ras protein expressions by using immunofluorescence assay. They found that the expression of P53 was dramatically up-regulated but the expression of H-Ras was hardly affected by safrole oxide, 224.72 micromol/L, within 24 hours. Taken together, these results revealed that safrole oxide could induce apoptosis of A549 cells and suggested that safrole oxide might perform its function by blocking cells completely at G1 phase and partly at G(2)-M phase, and also by up-regulating the expression of P53 protein. These findings would raise exciting possibilities for cancer therapy in future.     

肉桂酸对人肺腺癌A549细胞相关基因表达的影响

目的探讨肉桂酸（CINN）诱导肺癌A549细胞分化的能力及其分子机制。方法采用原位杂交和完整细胞原位斑点印迹等技术,研究CINN对人肺腺癌A549细胞分化相关蛋白质分子CD15,相关基因c-myc、EGFR、wtp53、wtp16等表达的影响。结果CINN可上调wtp53、wtp16基因,抑制CD15、c-myc、EGFR基因表达。结论CINN的上述作用可能是其诱导肺癌A549细胞分化的机制之一。     

Epithelial mesenchymal transition of non-small-cell lung cancer cells A549 induced by SPHK1

Objective:To explore the effect and molecular mechanism of SPHK1 in the invasion and metastasis process of non-small-cell lung cancer cells(A549).Methods:Recombinant retrovirus was used to mediate the production of A549/vector,A549/SPHK1,A549/scramble,and A549/SPHK1/RNAi that stably expressed or silenced SPHK1.The invasion and migration capacities of A549 cells overexpressing or silencing SPHK1 were determined using Transwell invasion assay and scratch wound repair experiment.The protein and mRNA expression levels of E-cadherin,fibronectin,vimentin in A549/vector,A549/SPHK1,A549/scramble,A549/SPHK1/RNAi were detected with Western blot(WB) and quantitative PCR(QPCR) methods,respectively.Results:Transwell invasion assay and scratch wound repair experiments showed that over-expression of SPHK1 obviously enhanced the invasion and migration capacities of A549 cells.WB and QPCR detection results showed that,the expression of E-cadherin(a molecular marker of epithelial cells) and fibronectin,vimentin(molecular markers of mesenchymal cells) in A549 cells was upregulated after overexpression of SPHK1;while SPHK1 silencing significantly reduced the invasion and metastasis capacities of A549 cells,upregulated the expression of molecular marker of epithelial cells,and downregulated the expression of molecular marker of mesenchymal cells.Conclusions:SPHK1 promotes epithelial mesenchymal transition of non-small-cell lung cancer cells and affects the invasion and metastasis capacities of these cells.     

人肺腺癌A549细胞系培美曲塞耐药细胞株模型的建立

目的建立人肺腺癌A549细胞培美曲塞耐药细胞株模型。方法以人肺腺癌A549细胞为亲本株,应用高浓度反复间歇法模拟临床给药模式建立人肺腺癌A549细胞培美曲塞耐药细胞株。MTT法绘制生长曲线、计算倍增时间,同时检测耐药株细胞对顺铂、依托泊苷、紫杉醇、长春新碱的药物敏感性。结果亲本株IC_50为(181.75±12.56)ng/ml,耐药株IC_50为(4930.40±129.71)ng/ml,最终获得耐药指数为27.18±1.16的人肺腺癌A549细胞培美曲塞耐药细胞株,命名为A549/PEM。A549和A549/PEM增殖速度接近,倍增时间分别为(62.78±1.06)h和(62.35±0.89)h(P=0.773)。A549/PEM同时对顺铂、依托泊苷耐药,耐药指数分别为(1.93±0.02)、(8.26±1.55);对紫杉醇、长春新碱敏感,耐药指数分别为(0.89±0.10)、(0.95±0.11)。结论本研究成功建立的人肺腺癌A549细胞培美曲塞耐药细胞株模型。     

Protein C inhibits endocytosis of thrombin-thrombomodulin complexes in A549 lung cancer cells and human umbilical vein endothelial cells

We investigated the effect of protein C on the endocytosis of thrombin- thrombomodulin complexes. We previously showed that exposure of umbilical vein endothelial cells to thrombin stimulated the internalization and degradation of thrombin. A similar internalization was stimulated by a monoclonal antithrombomodulin antibody. We have repeated these studies in the presence of protein C and found that endocytosis of 125I-thrombin-thrombomodulin complexes, but not 125I- antithrombomodulin-thrombomodulin complexes, is inhibited. Activated protein C did not inhibit endocytosis of thrombin-thrombomodulin complexes. Protein C inhibited both internalization and degradation of 125I-thrombin and diisopropylphosphoryl (DIP) 125I-thrombin in human lung cancer cells (A549). These effects were observed at protein C concentrations found in human plasma. Protein S had no effect on the inhibition of endocytosis of thrombin-thrombomodulin complexes by protein C. We propose that protein C may regulate the rate of endocytosis of thrombin-thrombomodulin complexes in vivo and thereby control the capacity for endothelium to activate protein C.     

Biosynthesis of calcitonin by human lung cancer cells

The ectopic secretion of calcitonin (CT) by a wide variety of nonthyroidal human tumors has been studied by CT RIA, but little information is available concerning the biosynthesis of CT in these tumors. In the present study, a human lung cancer cell line (BEN), secreting high mol wt forms of CT was investigated to characterize the CT gene products synthesized. When conditioned medium from BEN cells was chromatographed through a Bio-Gel P-30 column, larger species of immunoreactive CT were detected with mol wt of approximately 8,000 and 18,000. Little, if any, CT of 3,500 mol wt was detected. To examine CT gene products produced in BEN cells, poly A+ RNA was isolated from BEN cells and subjected to cell-free translation assays and DNA/RNA hybridization assays. In the wheat germ cell-free translation assay, a single BEN cell product which migrated on sodium dodecyl sulfate-polyacrylamide gels with an apparent mol wt of 17,000 could be specifically immunoprecipitated with CT antisera. A similar sized CT-related translation product is produced in wheat germ assays programmed by mRNA prepared from human medullary thyroid carcinomas. In DNA/RNA hybridization assays, a single BEN cell mRNA species of 1,000 base pairs, identical in size to human thyroidal CT mRNA, hybridized to a radiolabeled CT cDNA probe. Hybridization of the CT cDNA probe with BEN cell mRNA was confirmed by RNA dot blot hybridization and cytoplasmic RNA blotting procedures. These results indicate that larger mol wt forms of CT secreted by BEN cells are derived from a translation product and a mRNA which are of similar, if not identical, size as CT gene products produced in human thyroidal tissues. The inability of lung tumor cells to process the CT precursor to calcitonin of 3,500 mol wt may reflect a lack of specific prohormone processing enzymes in these tumor cells or may be due to structural polymorphism in the CT precursor expressed in the lung cells.     

蝙蝠葛活性成分对人肺癌A549细胞株抗增殖作用的实验研究

目的探讨蝙蝠葛活性成分对人肺癌A549细胞株抗增殖作用及其机制。方法应用MTT法测定蝙蝠葛活性成分对人肺癌A549细胞株的生长抑制作用;通过吖碇橙（AO）/溴化乙啶（EB）染色荧光显微镜观察肿瘤细胞的形态学变化;采用流式细胞仪检测A549细胞的周期分布相;应用免疫细胞化学技术SP法检测药物处理前后增殖细胞核抗原Ki-67、Bcl-2的表达。结果蝙蝠葛活性成分对人肺癌A549细胞株有明显的抑制生长的作用,且呈现出浓度依赖性;蝙蝠葛活性成分可诱导A549细胞发生细胞周期阻滞;蝙蝠葛活性成分作用后Ki-67和Bcl-2阳性表达率较对照组降低（P〈0.01）。结论蝙蝠葛活性成分在体外对人肺癌A549细胞株有显著的抑制增殖作用,可能与下调Ki-67、Bcl-2蛋白表达,细胞周期发生G0/G1期阻滞有关。     

Morphological observation of tumor infiltrating immunocytes in human rectal cancer

AIM: To investigate the morphological characterization of tumor infiltrating dendritic cells (TIDCs) and tumor infiltrating lymphocytes (TILs) in human rectal cancer. METHODS: Light and electron microscopy as well as im-munohistochemistry were used to observe the distributive and morphological changes of TIDCs and TILs. RESULTS: TIDCs were mainly located in tumor-surrounding tissue. The number of TIDCs in the earlier stage was higher than that in the later stage (P<0.01). TILs were mainly seen in adjacent tissue of cancers and tumor-surrounding tissue. There were more TILs in the earlier stage than that in the later stage (P<0.01). Under electron microscope, TIDCs were irregular in shape and exhibited many dendritic protrusions. It isn't obvious that cancer cells perforated the basement membrane and TILs were arranged along the basement membrane in the earlier stage. In the later stage, it is explicit that cancer cells perforated the basement membrane and surrounded by TILs. There were contacts among TIDCs, TILs and tumor cell. One TIDCs contacted one or several TILs which clustered around TIDCs. Glycogen granules were seen between TIDCs and TILs. CONCLUSION: The number of TIDCs and TILs is related with tumor progression There exist close relationships among TIDCs, TILs and tumor cell.